Chromosome segregation is a complex process requiring many molecular components. Improper chromosome segregation causes a variety of genetic disorders, including Downs syndrome. The proposed research is designed to identify and characterize genes encoding products involved in chromosome segregation in yeast. The goal of this research is to understand chromosome segregation at a molecular level. Four specific questions will be addressed: 1) Which genes encode products that are essential for proper chromosome segregation? Existing mutations will be identified by the specific requirement for their gene products at the time of chromosome segregation in the cell cycle. To isolate new chromosome-segregation mutations, we will exploit our recent observation that temperature-sensitive topoisomerase II mutations cause lethality if chromosome segregation proceeds at the restrictive temperature. 2) What do the phenotypes caused by mutations in these genes reveal about the process of chromosome segregation? The effects of the mutations on cell morphology, spindle morphology, nuclear morphology, and the progression of the cell division cycle will be examined. Perturbations of chromosome segregation leading to diploidization or chromosome instability will be determined genetically. The constellation of phenotypes caused by each mutation will provide information about the function of the gene product it identifies. 3) Which gene products interact physically with one another? A physical interaction between two gene products can be determined genetically by pseudoreversion analysis. A deleterious mutation in one protein can frequently be compensated by a mutation in an interacting protein. Such extragenic suppressors of chromosome-segregation mutations will be isolated, and they will be used to construct a map of physical interactions. 4) How do the gene products interact functionally? Cold-sensitive mutations will be combined genetically with temperature-sensitive mutations in other chromosome-segregation genes. After the double mutants are shifted from one restrictive temperature to the other, the success of chromosome segregation will be determined genetically and morphologically. The results of these experiments will reveal the order and dependency relationships of the steps identified by the mutations.